The Newcastle disease virus (NDV) belongs to serotype 1 of avian paramyxovirus belonging to the genus Avulavirus of the family Paramyxoviridae (Mayo, 2002). This disease is considered one of the pathological processes with greater economic impact on the poultry industry. The virus is capable to affect respiratory, digestive and nervous systems. Digestive tract secretions are a major source of infection for virus replication. The Newcastle disease virus is transmitted during the incubation period that lasts 4 to 6 days and for a limited period of time during convalescence. The severity of the disease symptoms depends on the genotype, strain, dose, immune status of the bird, exposition route, the presence of other infectious agents and environmental conditions. In acute cases, the only clinical manifestation of the disease is mortality.
To date, the presence of this disease is of great concern although there is no evidence of significant mortality problems probably because of the protection offered by vaccination. However, vaccination does not prevent viral shedding and dissemination.
It is essential to determine the pathogenicity of a strain based on its biological behavior and genetic structure.
According to its pathogenicity, the Newcastle disease virus is classified into three types: a) lentogenic, presenting low pathogenicity, b) mesogenic, presenting moderate pathogenicity, and c) velogenic, presenting high pathogenicity. At genetic level, the pathogenicity is mainly determined by the amino acid sequence present at the cleavage site of the fusion protein of the Newcastle virus, specifically the 5 amino acids before the cleavage site. Greater the number of basic amino acids, greater is the degree of pathogenicity. Thus, the velogenic phenotype corresponds to the presence of 4 basic amino acids while the lentogenic phenotype corresponds to the presence of two basic amino acids at the cleavage site. (Glickman et al. 1988; Peeters B., et al. 1999).
Six encoded proteins participating in the infection and replication are found in the virus genome: 1) a nucleoprotein (NP), the phosphoprotein, (P), the matrix protein (M), the fusion protein (F), an hemagglutinin-neuraminidase (HN) and an RNA-dependent RNA polymerase (L) (de Leeuw and Peeters, 1999).
Among the proteins encoded in the virus genome, two are the most important from an antigenic point of view: hemagglutinin-neuraminidase (HN) and the fusion protein (F). These two proteins found on the surface of the viral envelope are the target of the humoral immune response.
The function of the fusion protein is to perform the cell entry process of the viral particle. This protein is the main responsible for the pathogenicity phenotype. This phenomenon depends on the chemical properties of the amino acids present at the cleavage side of the protein. The predominance of basic amino acids expresses a velogenic phenotype, while their low presence characterizes lentogenic strains (Glickman et al. 1998).
The encoding gene for the HN protein is responsible for the activities related to hemagglutination and neuraminidase. Its main function is to promote cell binding, and the breaking of sialic acid chains, respectively. Immunologically, it is the most important of all the proteins of this virus because it is the most antigenic. The encoding gene for the protein responsible for the transcription of the viral genes and the replication (Cobaleda et al., 2002).
Studies conducted in the year 2000 by Dr. Swayne and Dr. Suarez showed that there are differences regarding the capacity to neutralize virus replication at respiratory tract level depending on the homology level existing between the vaccine virus and the challenge virus in a study performed with various high pathogenicity viruses. The presence of antigenic and genetic diversity with regard to Newcastle disease virus has been recognized and it is known that any strain can protect against the mortality caused by a challenge from another lineage but however epidemiological outbreaks continue to appear.
In studies conducted by Pedersen J C et al. (J. Clin Microbiol 2004, 42:2329-2334), it was shown by phylogenetic analyses that the viruses isolated in California, Nevada and Texas were related to viruses isolated in Mexico and Central America, that are sequentially remote from the La Sota strain isolated in 1946 in the United States of America, the strain used in most commercial vaccines.
Commercial vaccines are prepared using a lentogenic strain and are marketed as live virus and inactivated virus, being the La Sota strain of the Newscastle disease virus the most used worldwide, among others. This strain belongs to the genotype II, according to the classification proposed by Czegledi et al. in 2006.
Miller et al., 2007 (Vaccine 2007, 25: 7238-7246) report that the homologous vaccination with the challenge reduces virus elimination compared to the reduction observed in vaccines prepared with heterologous strains, facilitating viral circulation and therefore increasing the possibility of mutations, confirming thus the need to effectively reduce viral shedding for a better control of the disease.
Several technologies have been proposed to tackle these problems, for example, US application 2010/0183664 offers the use of nucleotides sequence encoding for NP, P, M and L proteins of a low pathogenicity virus and for F and HN proteins from a high pathogenicity virus from genotype VII, however it is considered that said vaccine may result in the emergence of new virus strains, and thus it is not appropriate for use as live vaccine.
U.S. Pat. No. 7,442,379 proposes the use of an RNA molecule comprising specific binding sites for a NCD virus polymerase and a mutation in the signals required to inhibit replication and transcription of a NCD virus linked to a RNA sequence derived from heterologous viruses such as the human immunodeficiency virus, Marek's disease, influenza, etc.
However, current vaccines are considered as a factor in the outbreak of diseases because there is no appropriately control of elimination of the virus, being necessary vaccination that should not only prevent mortality but also reduce viral shedding and eliminate or prevent mutations, i.e. it is necessary to have highly antigenic fragment vaccines.
Sequences List.
SEQ ID NO 1 corresponds to sequence fragment of the genome of the Newcastle disease virus encoding for F and HN proteins designed by phylogenetic analysis and identified as SEQ 1.
SEQ 2 corresponds to sequence of the Newcastle disease virus identified as APMV1/Chicken/Mexico/RecP05/2005 designed by phylogenetic analysis of strain APMV1/Chicken/Mexico/P05/2005 identified as SEQ 2.
SEQ 3 corresponds to sequence of the Newcastle disease virus identified as 1083 (Fontana)/72/RecP05 designed by phylogenetic analysis of strain 1083 (Fontana)/72 identified as SEQ 3
SEQ 4 corresponds to sequence of the Newcastle disease virus identified as Gamefowl/U.S.(CA)/211472/02/RecP05 designed by phylogenetic analysis of strain Gamefowl/U.S.(CA)/211472/02 identified as SEQ 4
SEQ 5 corresponds to sequence of the Newcastle disease virus identified as RecLSP05 designed by phylogenetic analysis of strain LaSota identified as SEQ 5.